Autoreactive CD4 and CD8 T cells and large numbers of B220+ CD4-CD8-double negative (DN) (( T cells accumulate in mice with impaired Fas-mediated apoptosis leading to autoimmune lymphoproliferation. Similar autoimmune disease (ALPS) occurs in humans with impaired Fas pathway. Paradoxically, T cell lymphoproliferation associated with loss of function in the Fas pathway does not result in overt T cell-mediated autoimmunity. Impairment of the Fas pathway even confers resistance to autoimmune diabetes in mice. Neither the function of B220+ DN T cells, nor the mechanism by which the autoreactive T cells in affected are controlled is clearly understood. We have examined and found that B220+ DN T cells are immunoregulatory T cells that suppress polyclonal and antigen specific T cell activation in vitro and prevent T cell-mediated colitis in an animal model of inflammatory bowel disease. In normal animals, B220+ DN T cells are found in the appendix and large intestine. We hypothesize that B220+ DN T cells are important for maintenance of mucosal and peripheral tolerance. We propose two specific aims to investigate this hypothesis: 1) To understanding the molecular mechanism of B220+ DN T cell-mediated suppression, we will use DNA microarray assay and in vitro and in vivo functional studies to identify genes whose products are involved in mediating B220+ DN T cell suppressor function. 2) To define the role of B220+ DN T cells in systemic and mucosal tolerance, we will analyze whether neonatal transfer of B220+ DN T cells prevents a) the fatal lymphoproliferation in scurfy mice or b) enterocolitis in IL-10 deficient mice. The findings generated by these studies will aid in characterizing a novel naturally occurring regulatory T cells and define their role in peripheral and mucosal tolerance. [unreadable] [unreadable]